CN108179189A - A kind of multiplex PCR specific primer and method for detecting TP53 gene mutations - Google Patents
A kind of multiplex PCR specific primer and method for detecting TP53 gene mutations Download PDFInfo
- Publication number
- CN108179189A CN108179189A CN201810061976.8A CN201810061976A CN108179189A CN 108179189 A CN108179189 A CN 108179189A CN 201810061976 A CN201810061976 A CN 201810061976A CN 108179189 A CN108179189 A CN 108179189A
- Authority
- CN
- China
- Prior art keywords
- primer
- gene
- library
- blood disease
- gene mutations
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
- C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
- C12Q1/686—Polymerase chain reaction [PCR]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6869—Methods for sequencing
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Analytical Chemistry (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- General Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Biophysics (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Microbiology (AREA)
- General Health & Medical Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Pathology (AREA)
- Hospice & Palliative Care (AREA)
- Oncology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The present invention relates to a kind of multiplex PCR specific primers and method for detecting TP53 gene mutations, more particularly to a kind of multiplex PCR specific primer and detection method based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations, multiple sample TP53 gene mutations can be detected simultaneously.The present invention breaks through the limitation of traditional sensing techniques using high-throughput gene sequencing technology, improves detection sensitivity；The variation situation including known mutations and unknown mutation site can also be detected simultaneously, obtains comprehensive and accurate testing result.By the way that TP53 gene mutation sites are sequenced, specific gene sequence information are can obtain, and then patient is subjected to prognosis parting, different therapeutic schemes is made according to different partings, is accomplished because of people's dispenser.
Description
Technical field
The invention belongs to biotechnologies, and in particular to one kind is related based on high throughput sequencing technologies detection blood disease
Multiplex PCR specific primer, method and the purposes of TP53 gene mutations.
Background technology
Hematological system tumor has become one of principal disease for threatening human life, and malignant hematologic disease is a kind of intractable blood
Liquid system tumor has the features such as treatment is intractable, and life cycle is short, poor prognosis, and pathogenesis is not yet completely understood so far.Right
In the treatment of malignant hematologic disease, good assessment is early diagnosed and reaches if can be made to disease, for improving facing for patient
Bed remission rate and prognosis evaluation extend efficient help, and improve existence and cure rate.
Gene-correlation research is constantly in leading positioning in cancer, and the detection of blood disease related gene is also relatively early enters
Clinical practice.In recent years, due to the development of Protocols in Molecular Biology, the research that changes to blood disease cellular elements science of heredity
It deepens continuously, it has been found that, there is chromosomal structural aberration in most blood disease, including lacking, repeating, inversion, easily
Position etc., leads to proto-oncogene and tumor suppressor gene structure variation so that protooncogene activation or tumor suppressor gene inactivation.When gene occurs
Variation, directly affects downstream signaling pathway, leads to ability of cell proliferation enhancing, apoptosis obstacle, dysdifferentiation etc., generates
Blood disease phenotype.With deep and technique of gene detection the development of blood disease pathogenesis, to related genes variants feelings
The detection of condition, the gradually foundation as blood disease diagnosis and treatment and reference.
TP53 genes are to study one of more important tumor suppressor gene at present, it is normal cell proliferation and breaks up negative
To regulatory factor, the tumor suppressor protein p53 of coding can make polygenes keep stability, and adjust the growth of cell, differentiation,
Aging avoids disease from generating, and is referred to as " gene defender ".When TP53 gene delections, mutation and disorderly regulation and control, by direct shadow
The function and activity of p53 albumen are rung, is occurred so as to cause tumour.Clinical research confirmation in recent years, the mutation status of TP53 genes
There is correlation with neoplastic hematologic disorder progression of disease, therapeutic effect, have in different blood disease hypotypes, different documents relatively uniform
Clinical meaning prompts prognosis mala.The guide for there was only MDS before 2016 has carried out specific prognosis mala point to TP53 mutation
Layer, the guide of multiple blood disease hypotypes is introduced sequentially into clinical meaning of the TP53 mutation to disease after 2016.Especially NCCN《It is anxious
Acute myeloid leukemia clinical practice guideline》, explicitly point out the patient that generation TP53 is mutated and belong to high-risk group.The AML of this case
Patient detects high-frequency TP53 mutation after the treatment, and indication curative effect may be bad.To sum up, the mutation status of TP53 genes is
Judge the important indicator of blood disease prognosis, be mutated using TP53 as tumor markers, neoplastic hematologic disorder is detected in real time, is passed through
Repeatedly sampling sequencing detection gene mutation situation determines that prognosis is layered, and gives clinical certain diagnosis and treatment guidance.In addition, the detection
The pain that patient operates on can be reduced, it is only necessary to which peripheral blood or marrow, sampling are convenient so that the state of an illness of patients with hematological tumor with
It visits lighter.
TP53 gene overall lengths are about 20kb, and clinical research at present does not sum up the mutantional hotspot of the gene, it has been reported that
The TP53 gene mutations of clinical meaning only detection can't have limit point instantly, need big model in each exon 1 of gene
Enclose detection.Therefore it in clinical practice work, needs to carry out sequencing analysis to the full extron of the gene to assist prognosis.Tradition
Detection method of gene mutation has Sanger methods, gene chips, fluorescence quantitative PCR method etc., these methods there are certain defect,
If the sequencing of Sanger PCR sequencing PCRs every time can only be sequenced a certain section of region of a sample, detection efficiency is low, of high cost, and
Sensitivity.Gene chips, fluorescence quantitative PCR method can only be directed to the site design detection probe shape of one or several known mutations
Into detection chip, therefore detection unknown mutation site is not used to, the result of detection is not comprehensive enough, accurate.Therefore, it is necessary to it sets
High throughput, highly sensitive detection method of the meter for TP53 gene mutations.
Invention content
It dashes forward in view of the above-mentioned problems, the present invention provides a kind of high throughput sequencing technologies detection blood disease correlation TP53 genes that are based on
The multiplex PCR specific primer and detection method of change, can detect multiple sample TP53 gene mutations simultaneously, and this method can be effective
Improve detection efficiency, accuracy rate, high sensitivity, high specificity.
To reach the object of the invention, using following technical scheme：
In a first aspect, a kind of multiplex PCR based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations is special
The design method of specific primer, comprises the following steps：
(1) the primer library in design object region；
(2) primer in the primer library of step (1) design is screened；
(3) specific primer that screening obtains adds universal joint to get amplimer to the end.
Preferably, the primer designed in step (1) covers all exon regions of TP53 genes.
Preferably, the primer screening principle in step (2) is：
(a) primer specificity is strong；
(b) not comprising simple repeated sequence；
(c) hot spot mutation position is avoided；
(d) it is compared to each other between primer so that interact between primer minimum；
It is further preferred that the interaction minimum between primer described in step (d) refers to before 3 ' ends of primer 8
There is no base pair complementarity in base, the continuous pairing base between primer and primer is no more than 5, whole complementary pairings
Base number be no more than the primer length 30%.
Preferably, such as SEQ ID NO of the universal joint primer sequence described in step (3)：29 and SEQ ID NO：30 institutes
Show, wherein, the end of sense primer 5 ' connection SEQ ID NO：29, the end of downstream primer 5 ' connection SEQ ID NO：30.
According to mentioned above principle, 14 pairs of specific primers, nucleotide sequence such as SEQ ID NO are finally obtained：1 to SEQ ID
NO：Shown in 28.
Second aspect, a kind of method based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations, comprising
Following steps：
(I) sample process and DNA extractions；
(II) PCR reaction systems and reaction condition are established, and is expanded using the primer pair destination region in step (3)
Increase；
(III) high-throughput library construction is carried out to the PCR product in step (II), and high pass measurement is carried out to the library
Sequence；
(IV) sequencing data and reference gene sequence are compared by analysis by sequence alignment program, obtained in patient body
The abrupt information of TP53 genes.
Preferably, the sample described in step (I) can be peripheral blood and/or marrow etc..
Preferably, the method for the extractions of DNA described in step (I), can be paramagnetic particle method and/or one kind in centrifugal column method or
Two kinds of combinations, currently preferred technical method are centrifugal column method.
In the present invention, the PCR reaction systems described in step (II) are as follows：
In the present invention, the PCR reaction conditions described in step (II) are as follows：
Preferably, each primer of primer mixture using concentration is TP53 according to existing document report in reaction system
Catastrophe of the gene in blood disease determines the final concentration of each primer in the reaction system.
It is further preferred that catastrophe refers to the adduction of all mutational site frequencies in each extron.
In the present invention, the high-throughput library constructing method described in step (III) can be that PCR amplification method builds library, also may be used
To be that RNA isolation kit builds library.
Preferably, PCR amplification method builds library step and is：
1. the PCR product in step (II) is purified；
2. PCR amplification is carried out, sequencing primer of the primer in reaction system for illumina routines, every primer final concentration
For 2uM；
3. being purified to product, corresponding library is obtained.
Preferably, RNA isolation kit builds library step and is：
1) end is repaired, and 5 ' protruding terminus is carried out with filling-in, while 3 ' protruding terminus are carried out by the extension of complementary strand
Cutting, makes the duplex ends of DNA form flat end；
2) phosphorylation is carried out to the product in 1)；
3) P1 and A connectors are connected on product after the phosphorylation in 2) using DNA ligase, which is Ion
Torrent platform routine sequence measuring joints；
4) PCR is enriched with；
5) 4) product in is purified, and complete library construction.
In the present invention, the method for step (III) described high-flux sequence can be sequenced in synthesis.
It is further preferred that microarray dataset is including but not limited to Illumina HiSeq/MiSeq, Ion in synthesis
Torrent PGM/Proton/S5 etc..
The present invention breaks through the limitation of traditional sensing techniques using high-throughput gene sequencing technology, improves detection sensitivity；Together
When can also detect the variation situation including known mutations and unknown mutation site, obtain comprehensive and accurate testing result.By right
TP53 gene mutation sites are sequenced, and can obtain specific gene sequence information, and then patient is carried out prognosis parting, according to
Different partings make different therapeutic schemes, accomplish because of people's dispenser.
Description of the drawings
The present invention has drawings described below：
Fig. 1 detects the method flow diagram of TP53 gene mutations.
Fig. 2 PCR amplification method Library development flow schematic diagrames.
Fig. 3 RNA isolation kit Library development flow schematic diagrames.
Specific embodiment
The present invention is described in further detail below in conjunction with attached drawing 1-3.
As shown in Figure 1, a kind of high throughput sequencing technologies detection blood disease correlation TP53 genes that are based on of the present invention are dashed forward
The design method of the multiplex PCR specific primer of change, comprises the following steps：
(1) the primer library in design object region；
(2) primer in the primer library of step (1) design is screened；
(3) specific primer that screening obtains adds universal joint to get amplimer to the end.
Preferably, the primer designed in step (1) covers all exon regions of TP53 genes.
Preferably, the primer screening principle in step (2) is：
(a) primer specificity is strong；
(b) not comprising simple repeated sequence；
(c) hot spot mutation position is avoided；
(d) it is compared to each other between primer so that interact between primer minimum；
It is further preferred that the interaction minimum between primer described in step (d) refers to before 3 ' ends of primer 8
There is no base pair complementarity in base, the continuous pairing base between primer and primer is no more than 5, whole complementary pairings
Base number be no more than the primer length 30%.
Preferably, such as SEQ ID NO of the universal joint primer sequence described in step (3)：29 and SEQ ID NO：30 institutes
Show, wherein, the end of sense primer 5 ' connection SEQ ID NO：29, the end of downstream primer 5 ' connection SEQ ID NO：30.
According to mentioned above principle, 14 pairs of specific primers, nucleotide sequence such as SEQ ID NO are finally obtained：1 to SEQ ID
NO：Shown in 28.
A kind of method based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations, comprises the following steps：
(I) sample process and DNA extractions；
(II) PCR reaction systems and reaction condition are established, and is expanded using the primer pair destination region in step (3)
Increase；
(III) high-throughput library construction is carried out to the PCR product in step (II), and high pass measurement is carried out to the library
Sequence；
(IV) sequencing data and reference gene sequence are compared by analysis by sequence alignment program, obtained in patient body
The abrupt information of TP53 genes.
Preferably, the sample described in step (I) can be peripheral blood and/or marrow etc..
Preferably, the method for the extractions of DNA described in step (I), can be paramagnetic particle method and/or one kind in centrifugal column method or
Two kinds of combinations, currently preferred technical method are centrifugal column method.
In the present invention, the PCR reaction systems described in step (II) are as follows：
In the present invention, the PCR reaction conditions described in step (II) are as follows：
Preferably, each primer of primer mixture using concentration is TP53 according to existing document report in reaction system
Catastrophe of the gene in blood disease determines the final concentration of each primer in the reaction system.
It is further preferred that catastrophe refers to the adduction of all mutational site frequencies in each extron.
In the present invention, the high-throughput library constructing method described in step (III) can be that PCR amplification method builds library, also may be used
To be that RNA isolation kit builds library.
Preferably, PCR amplification method builds library step and is：
1. the PCR product in step (II) is purified；
2. PCR amplification is carried out, sequencing primer of the primer in reaction system for illumina routines, every primer final concentration
For 2uM；
3. being purified to product, corresponding library is obtained.
Preferably, RNA isolation kit builds library step and is：
1) end is repaired, and 5 ' protruding terminus is carried out with filling-in, while 3 ' protruding terminus are carried out by the extension of complementary strand
Cutting, makes the duplex ends of DNA form flat end；
2) phosphorylation is carried out to the product in 1)；
3) P1 and A connectors are connected on product after the phosphorylation in 2) using DNA ligase, which is Ion
Torrent platform routine sequence measuring joints；
4) PCR is enriched with；
5) 4) product in is purified, and complete library construction.
In the present invention, the method for step (III) described high-flux sequence can be sequenced in synthesis.
It is further preferred that microarray dataset is including but not limited to Illumina HiSeq/MiSeq, Ion in synthesis
Torrent PGM/Proton/S5 etc..
The content not being described in detail in this specification belongs to the prior art well known to professional and technical personnel in the field.
Claims (10)
1. a kind of multiplex PCR specific primer based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations is set
Meter method, it is characterised in that：It comprises the following steps：
(1) the primer library in design object region；
(2) primer in the primer library of step (1) design is screened；
(3) specific primer that screening obtains obtains amplimer to the end plus universal joint.
2. the multiplex PCR as described in claim 1 based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations
The design method of specific primer, it is characterised in that：The primer designed in step (1) covers all outer aobvious of TP53 genes
Subregion.
3. the multiplex PCR as described in claim 1 based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations
The design method of specific primer, it is characterised in that：Primer screening principle in step (2) is：
(a) primer specificity is strong；
(b) not comprising simple repeated sequence；
(c) hot spot mutation position is avoided；
(d) it is compared to each other between primer so that interact between primer minimum；
Interaction minimum between primer described in step (d) refers to that there is no bases in 8 bases before 3 ' ends of primer
Complementary pairing, the continuous pairing base between primer and primer are no more than 5, and the base number of whole complementary pairings is no more than
The 30% of the primer length.
4. the multiplex PCR as described in claim 1 based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations
The design method of specific primer, it is characterised in that：The primer sequence of universal joint such as SEQ ID NO described in step (3)：29
With SEQ ID NO：Shown in 30, wherein, the end of sense primer 5 ' connection SEQ ID NO：29, the end of downstream primer 5 ' connection SEQ ID
NO：30.
5. blood disease correlation TP53 genes are detected based on high throughput sequencing technologies as described in claim 1-4 any claims
The design method of the multiplex PCR specific primer of mutation, it is characterised in that：The amplimer includes 14 pairs of specific primers,
Its nucleotide sequence such as SEQ ID NO：1 to SEQ ID NO：Shown in 28.
A kind of 6. method based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations, it is characterised in that：Comprising
Following steps：
(I) sample process and DNA extractions；
(II) PCR reaction systems and reaction condition are established, and destination region is expanded using the amplimer in claim 5
Increase；
(III) high-throughput library construction is carried out to the PCR product in step (II), and high-flux sequence is carried out to the library；
(IV) sequencing data and reference gene sequence are compared by analysis by sequence alignment program, obtain TP53 in patient body
The abrupt information of gene.
7. the method as claimed in claim 6 based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations,
It is characterized in that：Sample described in step (I) is peripheral blood and/or marrow；The method of DNA extraction for paramagnetic particle method and/or
The combination of one or both of centrifugal column method.
8. the method as claimed in claim 6 based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations,
It is characterized in that：The use concentration of each primer of primer mixture is according to existing document report in PCR reaction systems in step (II)
Catastrophe of the TP53 genes in road in blood disease determines the final concentration of each primer in the reaction system；The catastrophe
Refer to the adduction of all mutational site frequencies in each extron.
9. the method as claimed in claim 6 based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations,
It is characterized in that：High-throughput library constructing method described in step (III) is that PCR amplification method builds library or RNA isolation kit builds library；
The PCR amplification method builds library step：
1. the PCR product in step (II) is purified；
2. carrying out PCR amplification, the primer in reaction system is the sequencing primer of illumina routines, and every primer is final concentration of
2uM；
3. being purified to product, corresponding library is obtained；
The RNA isolation kit builds library step：
1) end is repaired, and filling-in is carried out to 5 ' protruding terminus, while 3 ' protruding terminus are cut by the extension of complementary strand,
The duplex ends of DNA is made to form flat end；
2) phosphorylation is carried out to the product in 1)；
3) P1 and A connectors are connected on product after the phosphorylation in 2) using DNA ligase, which is Ion torrent
Platform routine sequence measuring joints；
4) PCR is enriched with；
5) 4) product in is purified, and complete library construction.
10. the method as claimed in claim 6 based on high throughput sequencing technologies detection blood disease correlation TP53 gene mutations,
It is characterized in that：The method of step (III) described high-flux sequence is to be sequenced in synthesis.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810061976.8A CN108179189A (en) | 2018-01-23 | 2018-01-23 | A kind of multiplex PCR specific primer and method for detecting TP53 gene mutations |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810061976.8A CN108179189A (en) | 2018-01-23 | 2018-01-23 | A kind of multiplex PCR specific primer and method for detecting TP53 gene mutations |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN108179189A true CN108179189A (en) | 2018-06-19 |
Family
ID=62551238
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810061976.8A Pending CN108179189A (en) | 2018-01-23 | 2018-01-23 | A kind of multiplex PCR specific primer and method for detecting TP53 gene mutations |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108179189A (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105624274A (en) * | 2014-11-06 | 2016-06-01 | 张煜 | High flux detection method for tumor-targeted drugs related genes mutation, primers and reagent thereof |
| CN106755418A (en) * | 2016-12-24 | 2017-05-31 | 广州艾基生物技术有限公司 | A kind of extron assembles sequence measurement |
| CN107447258A (en) * | 2016-06-01 | 2017-12-08 | 大连医科大学 | Circulating tumor DNA target gene high flux detects library and its detection method and application |
-
2018
- 2018-01-23 CN CN201810061976.8A patent/CN108179189A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105624274A (en) * | 2014-11-06 | 2016-06-01 | 张煜 | High flux detection method for tumor-targeted drugs related genes mutation, primers and reagent thereof |
| CN107447258A (en) * | 2016-06-01 | 2017-12-08 | 大连医科大学 | Circulating tumor DNA target gene high flux detects library and its detection method and application |
| CN106755418A (en) * | 2016-12-24 | 2017-05-31 | 广州艾基生物技术有限公司 | A kind of extron assembles sequence measurement |
Non-Patent Citations (3)
| Title |
|---|
| CLAUDIA VOLLBRECHT等: "Comprehensive Analysis of Disease-Related Genes in Chronic Lymphocytic Leukemia by Multiplex PCR-Based Next Generation Sequencing", 《POLS ONE》 * |
| TJ PUGH等: "Exome sequencing of pleuropulmonary blastoma reveals frequent biallelic loss of TP53 and two hits in DICER1 resulting in retention of 5p-derived miRNA hairpin loop sequences", 《ONCOGENE》 * |
| 李玉玲: "高通量测序技术在骨髓增殖性肿瘤临床检测中的方法学建立及其初步应用", 《中国优秀硕士学位论文全文数据库》 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN109207594B (en) | Method for detecting microsatellite stability state and genome change through plasma based on next generation sequencing | |
| CN108753967A (en) | A kind of gene set and its panel detection design methods for liver cancer detection | |
| CN104745679A (en) | Method and kit for non-invasive detection of EGFR (epidermal growth factor receptor) gene mutation | |
| CN105506746A (en) | Method for constructing variable region sequencing library, and method for determining variable region nucleic acid sequence | |
| US9663826B2 (en) | System and method of genomic profiling | |
| CN106755297A (en) | One group is based on primer sets that ARMS fluorescence quantitative PCR detection EGFR genes T790M is mutated and preparation method thereof | |
| WO2019076018A1 (en) | Method for constructing amplicon library for detecting low-frequency mutation of target gene | |
| CN106103742A (en) | A kind of method of enrichment cycles Tumour DNA and reagent | |
| CN106897579A (en) | New infantile tumour label and application based on chromosomal variation index | |
| CN110241215A (en) | A kind of primer, kit and detection method to make a variation for detecting Benign Thyroid Nodules tumor- associated gene | |
| CN108949979A (en) | A method of judging that Lung neoplasm is good pernicious by blood sample | |
| CN102399872B (en) | Method and kit for detecting mutation of K-ras gene | |
| CN110004229A (en) | Application of the polygenes as EGFR monoclonal antibody class Drug-resistant marker | |
| CN109735622A (en) | LncRNA relevant to colorectal cancer and its application | |
| CN107641649B (en) | Primer pair, kit and method for detecting stability of NR27 locus of microsatellite | |
| CN104480215A (en) | Gene joint detection method and kit | |
| CN103667267B (en) | For with the DNA probe storehouse of KRAS gene recombination and the method adopting its enrichment KRAS gene fragment | |
| CN108179189A (en) | A kind of multiplex PCR specific primer and method for detecting TP53 gene mutations | |
| CN107119122B (en) | Kit for detecting single nucleotide polymorphism and detection method | |
| CN102827935B (en) | Reagent kid for quantitatively testing mRNA (messenger ribonucleic acid) level of FIP1L1-PDGFRA (feline infectious peritonitis 1 like 1-platelet-derived growth factor receptor alpha) fusion genes | |
| CN102876799B (en) | Multiple real-time quantitative polymerase chain reaction (PCR) kit for detecting mixed lineage leukemia (MLL) relative fuse genes | |
| CN107586851A (en) | A kind of KRAS, NRAS, BRAF gene mutation detection kit | |
| CN108796072B (en) | Gene for molecular typing of hyper-mutant tumor and application thereof | |
| CN102586472B (en) | Method and kit for assaying hepatitis B virus DNA (deoxyribonucleic acid) sequence | |
| CN109811041A (en) | A kind of specific primer detecting the site PIK3CA gene H1047R is to, probe and kit |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20180619 |
|
| RJ01 | Rejection of invention patent application after publication |